Recherche de la fonction du récepteur orphelin couplé aux protéines G GPR50 par hétérodimérisation

LE KREMLIN-B.- PARIS 11-BU Méd (940432101) / SudocPARIS-BIUP (751062107) / SudocSudocFranceF

L’hétérodimérisation des récepteurs couplés aux protéines G

Les récepteurs couplés aux protéines G (RCPG) ou récepteurs à 7 domaines transmembranaires (7TM) représentent la plus grande famille de protéines de la surface cellulaire. Cette famille de protéines contrôle de nombreux processus physiologiques. Son implication dans de nombreuses pathologies fait de cette classe de protéines la cible d’environ 50 % des médicaments couramment prescrits aujourd’hui. Cependant, malgré les efforts réalisés par la recherche académique et l’industrie, une centaine de RCPG sont encore orphelins. Ces derniers présentent donc un intérêt particulier puisqu’ils constituent autant de cibles potentielles pour la découverte de nouvelles drogues. Les stratégies de recherche concernant les RCPG orphelins ont consisté essentiellement en l’identification de leurs ligands naturels. Tout comme dans le cas de certains récepteurs orphelins nucléaires, des données récentes ont montré que des RCPG orphelins peuvent réguler, par hétérodimérisation et indépendamment d’un ligand, la fonction d’autres récepteurs ayant un ligand connu. Cette caractéristique permettrait, d’une part, de mieux comprendre la diversité extraordinaire offerte par les RCPG et, d’autre part, d’ouvrir de nouvelles perspectives concernant l’identification de la fonction de ces RCPG orphelins, quels que soient les domaines thérapeutiques considérés

CXCR7 heterodimerizes with CXCR4 and regulates CXCL12-mediated G protein signaling.

International audienceThe stromal cell-derived factor-1/CXCL12 chemokine engages the CXCR4 and CXCR7 receptors and regulates homeostatic and pathologic processes, including organogenesis, leukocyte homeostasis, and tumorigenesis. Both receptors are widely expressed in mammalian cells, but how they cooperate to respond to CXCL12 is not well understood. Here, we show that CXCR7 per se does not trigger G(alphai) protein-dependent signaling, although energy transfer assays indicate that it constitutively interacts with G(alphai) proteins and undergoes CXCL12-mediated conformational changes. Moreover, when CXCR4 and CXCR7 are coexpressed, we show that receptor heterodimers form as efficiently as receptor homodimers, thus opening the possibility that CXCR4/CXCR7 heterodimer formation has consequences on CXCL12-mediated signals. Indeed, expression of CXCR7 induces conformational rearrangements within preassembled CXCR4/G(alphai) protein complexes and impairs CXCR4-promoted G(alphai)-protein activation and calcium responses. Varying CXCR7 expression levels and blocking CXCL12/CXCR7 interactions in primary T cells suggest that CXCR4/CXCR7 heterodimers form in primary lymphocytes and regulate CXCL12-promoted chemotaxis. Taken together, these results identify CXCR4/CXCR7 heterodimers as distinct functional units with novel properties, which can contribute to the functional plasticity of CXCL12

Do orphan G-protein-coupled receptors have ligand-independent functions? New insights from receptor heterodimers

G-protein-coupled receptors (GPCRs) are important drug targets and are involved in virtually every biological process. However, there are still more than 140 orphan GPCRs, and deciphering their function remains a priority for fundamental and clinical research. Research on orphan GPCRs has concentrated mainly on the identification of their natural ligands, whereas recent data suggest additional ligand-independent functions for these receptors. This emerging concept is connected with the observation that orphan GPCRs can heterodimerize with GPCRs that have identified ligands, and by so doing regulate the function of the latter. Pairing orphan GPCRs with their potential heterodimerization partners will have a major impact on our understanding of the extraordinary diversity offered by GPCR heterodimerization and, in addition, will constitute a novel strategy to elucidate the function of orphan receptors that needs to be added to the repertoire of ‘deorphanization' strategies

The PDZ Protein Mupp1 Promotes G i Coupling and Signaling of the Mt 1 Melatonin Receptor

International audienceIntracellular signaling events are often organized around PDZ (PSD-95/Drosophila Disc large/ZO-1 homology) domain-containing scaffolding proteins. The ubiquitously expressed multi-PDZ protein MUPP1, which is composed of 13 PDZ domains, has been shown to interact with multiple viral and cellular proteins and to play important roles in receptor targeting and trafficking. In this study, we show that MUPP1 binds to the G protein-coupled MT 1 melatonin receptor and directly regulates its G i-dependent signal transduction. Structural determinants involved in this interaction are the PDZ10 domain of MUPP1 and the valine of the canonical class III PDZ domain binding motif DSV of the MT 1 carboxyl terminus. This high affinity interaction (K d ϳ 4 nM), which is independent of MT 1 activation, occurs in the ovine pars tuberalis of the pituitary expressing both proteins endogenously. Although the disruption of the MT 1 /MUPP1 interaction has no effect on the subcel-lular localization, trafficking, or degradation of MT 1 , it destabilizes the interaction between MT 1 and G i and abolishes G i-mediated signaling of MT 1. Our findings highlight a previously unappreciated role of PDZ proteins in promoting G protein coupling to receptors

Leukocyte analysis from WHIM syndrome patients reveals a pivotal role for GRK3 in CXCR4 signaling.

International audienceLeukocytes from individuals with warts, hypogammaglobulinemia, infections, and myelokathexis (WHIM) syndrome, a rare immunodeficiency, and bearing a wild-type CXCR4 ORF (WHIM(WT)) display impaired CXCR4 internalization and desensitization upon exposure to CXCL12. The resulting enhanced CXCR4-dependent responses, including chemotaxis, probably impair leukocyte trafficking and account for the immunohematologic clinical manifestations of WHIM syndrome. We provided here evidence that GPCR kinase-3 (GRK3) specifically regulates CXCL12-promoted internalization and desensitization of CXCR4. GRK3-silenced control cells displayed altered CXCR4 attenuation and enhanced chemotaxis, as did WHIM(WT) cells. These findings identified GRK3 as a negative regulator of CXCL12-induced chemotaxis and as a candidate responsible for CXCR4 dysfunction in WHIM(WT) leukocytes. Consistent with this, we showed that GRK3 overexpression in both leukocytes and skin fibroblasts from 2 unrelated WHIM(WT) patients restored CXCL12-induced internalization and desensitization of CXCR4 and normalized chemotaxis. Moreover, we found in cells derived from one patient a profound and selective decrease in GRK3 products that probably resulted from defective mRNA synthesis. Taken together, these results have revealed a pivotal role for GRK3 in regulating CXCR4 attenuation and have provided a mechanistic link between the GRK3 pathway and the CXCR4-related WHIM(WT) disorder

High-Density Lipoprotein Therapy in Stroke: Evaluation of Endothelial SR-BI-Dependent Neuroprotective Effects

International audienceHigh-density lipoproteins (HDLs) display endothelial protective effects. We tested the role of SR-BI, an HDL receptor expressed by endothelial cells, in the neuroprotective effects of HDLs using an experimental model of acute ischemic stroke. After transient intraluminal middle cerebral artery occlusion (tMCAO), control and endothelial SR-BI deficient mice were intravenously injected by HDLs or saline. Infarct volume and blood-brain barrier (BBB) breakdown were assessed 24 h post tMCAO. The potential of HDLs and the role of SR-BI to maintain the BBB integrity was assessed by using a human cellular model of BBB (hCMEC/D3 cell line) subjected to oxygen-glucose deprivation (OGD). HDL therapy limited the infarct volume and the BBB leakage in control mice relative to saline injection. Interestingly, these neuroprotective effects were thwarted by the deletion of SR-BI in endothelial cells and preserved in mice deficient for SR-BI in myeloid cells. In vitro studies revealed that HDLs can preserve the integrity of the BBB in OGD conditions, and that this effect was reduced by the SR-BI inhibitor, BLT-1. The protection of BBB integrity plays a pivotal role in HDL therapy of acute ischemic stroke. Our results show that this effect is partially mediated by the HDL receptor, SR-BI expressed by endothelial cells

Differential activity and selectivity of N-terminal modified CXCL12 chemokines at the CXCR4 and ACKR3 receptors

International audienceChemokines play critical roles in numerous physiological and pathological processes through their action on seven-transmembrane (TM) receptors. The N-terminal domain of chemokines, which is a key determinant of signaling via its binding within a pocket formed by receptors' TM helices, can be the target of proteolytic processing. An illustrative case of this regulatory mechanism is the natural processing of CXCL12 that generates chemokine variants lacking the first two N-terminal residues. While such truncated variants behave as antagonists of CXCR4, the canonical G proteincoupled receptor of CXCL12, they are agonists of the atypical chemokine receptor 3 (ACKR3